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Onconase and amphinase, the antitumor ribonucleases from Rana pipiens oocytes
W Ardelt1, K Shogen, Z Darzynkiewicz
1Alfacell Corporation, 300 Atrium Drive, Somerset, NJ 08873, USA. wardelt@alfacell.com
Abstract:
Rana pipiens oocytes contain two homologues of pancreatic ribonuclease A that are cytostatic and cytotoxic to human cancer cells. Extensively studied Onconase is in advanced Phase IIIb clinical trials against malignant mesothelioma, while Amphinase is a novel enzyme in pre-clinical development. Onconase is the smallest (104 amino acid residues) member of the ribonuclease A superfamily while Amphinase (114 residues) is the largest among amphibian ribonucleases. Both enzymes share the characteristic frog ribonucleases C-terminal disulfide bond but another signature of this group, the N-terminal pyroglutamate, an integral part of Onconase active site is not conserved in Amphinase. Although Onconase and Amphinase are weak catalysts their enzymatic activities are required for cytostatic and cytotoxic activity. While it was postulated that tRNA is the primary substrate of Onconase in vivo there is also extensive indirect evidence that suggests other RNA species, in particular micro RNAs, may actually be the critical target of these ribonucleases. The cytostatic effects of Onconase and Amphinase are manifested as cell arrest in the G(1) cell cycle phase. Apoptosis then follows involving activation of endonucleases(s), caspases, serine proteases and transglutaminase. Onconase was shown to be strongly synergistic when combined with numerous other antitumor modalities. Onconase and Amphinase are highly cationic molecules and their preferential toxicity towards cancer cells (having distinctly higher negative charge compared to normal cells) may depend on increased binding efficiency to the cell surface by electrostatic interactions.
Insights
Two frog-derived ribonucleases, Onconase and Amphinase, show potent anticancer activity by targeting RNA in cancer cells. Their unique properties and mechanisms offer promising therapeutic potential for various cancers.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Rana pipiens oocytes yield two pancreatic ribonuclease A homologues with demonstrated cytostatic and cytotoxic effects on human cancer cells.
- Onconase, a small ribonuclease, is in Phase IIIb trials for malignant mesothelioma; Amphinase, a larger homologue, is in pre-clinical development.
Purpose of the Study:
- To investigate the structural and functional characteristics of Onconase and Amphinase.
- To elucidate the mechanism of action and therapeutic potential of these ribonucleases against cancer cells.
Main Methods:
- Comparative analysis of enzyme structure (amino acid sequence, disulfide bonds, N-terminal pyroglutamate).
- Assessment of enzymatic activity and its correlation with cytostatic/cytotoxic effects.
- Investigation of cellular targets (tRNA, microRNAs) and cell cycle effects (G1 arrest, apoptosis).
- Evaluation of synergistic effects with other anti-cancer agents and the role of cationic properties in cancer cell targeting.
Main Results:
- Onconase (104 residues) and Amphinase (114 residues) possess distinct structural features, including the conserved C-terminal disulfide bond but differing N-terminal pyroglutamate.
- Enzymatic activity, though weak, is crucial for their anti-cancer effects, potentially targeting microRNAs.
- Both enzymes induce G1 cell cycle arrest and apoptosis, with Onconase showing synergy with other therapies.
- High cationic nature facilitates preferential binding and toxicity towards negatively charged cancer cells.
Conclusions:
- Onconase and Amphinase are potent anti-cancer agents derived from amphibian ribonucleases.
- Their efficacy relies on specific structural elements, enzymatic activity against RNA targets, and electrostatic interactions with cancer cells.
- These enzymes represent promising candidates for novel cancer therapeutics, with Onconase already in advanced clinical trials.
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